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Journal of Virology, April 2007, p. 4272-4285, Vol. 81, No. 8
0022-538X/07/$08.00+0     doi:10.1128/JVI.02536-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Antigenic and Immunogenic Study of Membrane-Proximal External Region-Grafted gp120 Antigens by a DNA Prime-Protein Boost Immunization Strategy{triangledown} ,{dagger}

Mansun Law,1 Rosa M. F. Cardoso,2 Ian A. Wilson,2 and Dennis R. Burton1,2*

Departments of Immunology,1 Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 920372

Received 16 November 2006/ Accepted 19 January 2007

The membrane-proximal external region (MPER) of human immunodeficiency virus type 1 (HIV-1) gp41 is a target of broadly neutralizing monoclonal antibodies (MAbs) 2F5, 4E10, and Z13. Here we engrafted the MPER into the V1/2 region of HIV-1 gp120 to investigate the ability of the engineered antigens to elicit virus-neutralizing antibodies (NAbs). To promote the correct folding and presentation of the helical 4E10 epitope, we flanked the epitope with helical domains and manipulated the helix by sequential deletion of residues preceding the epitope. Binding of the recombinant proteins to MAb 4E10 increased four- to fivefold with the deletion of one or two residues, but it returned to the wild-type level when three residues were deleted, suggesting rotation of the 4E10 epitope along the helix. Immunization of mice and rabbits by electroporation-mediated DNA priming and protein boosting with these constructs elicited high levels of gp120-specific antibodies. A consistent NAb response against the neutralization-resistant, homologous JR-FL virus was detected in rabbits but not in mice. Analysis of the neutralizing activity revealed that the NAbs do not target the MPER or the V1, V2, or V3 region. Through this study, we learned the following. (i) The 4E10 epitope can be manipulated using a "rotate-the-helix" strategy that alters the helix register. However, presentation of this epitope in the immunogenic V1/2 region did not render it immunogenic in mice or rabbits. (ii) DNA vaccination with monomeric gp120-based antigens can elicit a consistent NAb response against the homologous neutralization-resistant virus by targeting epitopes outside the V1, V2, and V3 regions.

* Corresponding author. Mailing address: The Scripps Research Institute, Department of Immunology (IMM-2), 10550 North Torrey Pines Road, La Jolla, CA 92037. Phone: (858) 784-9298. Fax: (858) 784-8360. E-mail: burton{at}scripps.edu

{triangledown} Published ahead of print on 31 January 2007.

{dagger} This is article 18567 from The Scripps Research Institute.

Journal of Virology, April 2007, p. 4272-4285, Vol. 81, No. 8
0022-538X/07/$08.00+0     doi:10.1128/JVI.02536-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



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